Cell locomotion is fundamental to development, wound healing, aspects of immune system function and metastatic cancer. The general goal of this research has been to determine "what molecules move where" during cell movement in order to test existing models for locomotion. We propose to continue this effort, focusing on the relation of these movements to the functions required for locomotion. This research will compliment our increasing knowledge of the molecules involved in motility and lead to more sophisticated models for cell movement. The specific aims of this proposal are fourfold: (1) to determine directed movements of the plasma membrane constituents which accompany extensions of the leading edge and retraction of the trailing edge; 2) to determine the dynamics of focal and close contact formation and disassembly in terms of their constituent molecules as cells move on substrates; (3) to explore molecular factors which may regulate individual substratum contacts; and (4) to measure force development during retraction and other phenomena associated with locomotion in relation to substratum contact status. This work will make extensive use of recent developments in optical microscopy, used in conjunction with labeled antibodies to membrane components and fluorescent cytoskeletal analogs, to explore the dynamics of the plasma membrane and cell substratum contacts during cell movement. Digitized fluorescence microscopy, video fluorescence recovery after photobleaching, confocal microscopy, and colloidal gold particle tracking (Nanovid) structural regulators, will be employed. Rapid release of "caged" compounds suspected to be structural regulators, will be used to explore the control of cell- substratum contact structures. The bending of calibrated microneedles attached to the cell surface will be employed to measure forces involved in cell locomotion. It is anticipated that the information generated in this project will ultimately lead to a deepened understanding of the role of the cell movement in normal and pathological states.